1. Field of the Invention
This invent&on relates to an apparatus and a method for continually monitoring the presence liquids of toxic substances, bacteria or any substance or organism which leads to change in the luminescent output of luminescent organisms or luminescence systems. The method involves adding a substance or substances or organisms capable of emitting light to the liquid and analysing light emitted from the resulting mixture.
2. Discussion of Prior Art
Analysis by emitted light is an effective and rapid method for hygiene monitoring and toxicity testing. Rapid and highly sensitive methods for toxicity testing have been based upon the use of luminescent bacteria, in which a reduction of the light emitted by the bacteria is proportional to the presence of toxic materials in the sample. Such tests are extremely sensitive and quantitative. They are used for example by manufacturers in quality assurance of raw materials and to monitor quality during manufacture. They are used to measure the toxicity of untreated and treated wastewaters and as an alternative to animal testing by the cosmetics industry. Their widespread use is being encouraged in the U.K. by the National Rivers Authority.
A number of commercially available luminescence tests for hygiene and toxicity monitoring are currently available and are either based on the luminescent system of the firefly or that of a number of species of luminescent bacteria. They are all performed manually and individually on previously collected samples. They could not be relied upon to monitor unexpected episodes in which pollutants are deliberately or accidentally discharged into a water course. They could not be used easily to monitor changes in the quality of a liquid in a process plant or in the continual quality control of washing of products. They are severely limited by the number of samples that can be collected and the interval between samples. Using an autosampler could increase the number of samples analysed but the corresponding increased cost in reagents would limit the popularity of such an automated system.
U.S. 4,357,420 discloses a method for the detection of biomarkers from biological fluids using luminescence. This process involves the chromatographic separation of samples of biological fluids in which specific biomarkers may be detected. The method uses individual samples of biological fluids and is unsuitable for the detection of unknown biomarkers.
GB 2005018 discloses a method for detecting a toxic substance by detecting the change in light output from a suspension of luminescent microorganisms in an aqueous liquid. The method is, however, based on the analysis of discrete samples of liquid.
U.S. 4,385,113 teaches the use of a bioluminescent system to assay for the presence of ATP and, hence, microorganisms in an aqueous system.
Bains, in Biotechnology, volume 10, May 1992, pages 515 to 518, describes commercially available sensors and states that since they are not continuous sensors, they have found only limited use.
Previously known systems rely on providing a reservoir of bacterial culture either freshly grown in a closed batch culture system or reconstituted from dried bacterial culture but ensuring in some way that the bacteria do not multiply.
Luminescent bacteria grown in closed culture will exhibit various stages of growth:lag phase, acceleration phase, exponential phase, retardation phase and death phase. Closed batch culture vessels will contain a mixture of microorganisms at various stages of the cell cycle. The number of microorganisms will increase at varying rates during the different phases of growth of a culture and the culture will eventually decline and die. As the number of organisms in the culture increases the environment in which they live changes. Many cellular components, such as ATP, DNA and proteins alter in response to these changes and thus the differences between individuals in their biochemical responses (such as luminescence) will also vary. It is for many of the above reasons that the currently used batch tests are designed to prevent the growth of the fresh or reconstituted dried bacteria.
There is however another way to grow microorganisms and that is in open growth systems where there is a continuous input of growth substrates (medium) and removal of waste products, cells and unused substrate. Parameters such as pH, oxygen, temperature etc. can be monitored during the growth with any changes automatically compensated for, for example by the addition of substances to reduce or increase the pH, change the temperature, oxygen content or other conditions.
In these continuous flow cultures the exponential growth phase is prolonged indefinitely as additions to and removal from the culture take place continuously. Thus the characteristics of the population of microorganisms, and that part of it removed for use, remain constant.